AOAC-RI ERP Book MICRO Sept 2016.pdf

B ird et al .: J ournal of AOAC I nternational V ol . 97, N o . 6, 2014  1565

Both test portion sizes analyzed by the 3M Petrifilm SALX System were compared to samples (25 g) analyzed using either the MLG or BAM reference method in an unpaired study design. All positive test portions were biochemically confirmed by the API 20E biochemical test, AOAC Official Method 978.24 or by the VITEK 2 GN identification test, AOAC Official Method 2011.17 . The biochemical method was determined by each individual participating laboratory based on their current method used for confirmation of routine samples. Serological testing, Group Poly OA-I & Vi and Poly H latex agglutination, was also performed. Each collaborating laboratory recorded results for the reference method and the 3M Petrifilm SALX System on the data sheets provided in the collaborative study outline or the electronic spreadsheet created as a result of multiple requests for electronic data entry. The data sheets were submitted to the Study Director at the end of each week of testing for analysis. The results of each test portion for each sample were compiled by the Study Director and the qualitative 3M Petrifilm SALX System results were compared to the reference methods for statistical analysis. Data for each test portion size were analyzed using the probability of detection (POD) statistical model (3, 7). A confidence interval of a dLPOD (difference between the POD of the reference and candidate method) not containing the point zero would indicate a statistically significant difference between the 3M Petrifilm SALX System and the MLG or BAM reference methods at the 5% probability level (8). In addition to calculating the POD for each inoculation level, the repeatability standard deviation, among-laboratory standard deviation, reproducibility standard deviation, and a P -value for homogeneity were calculated. For the collaborative study, the 3M Petrifilm SALX System produced 479 presumptive positive results with 475 confirming positive by the traditional confirmation and 473 confirming positive by the alternative confirmation. There were 468 confirmed positives by the reference method. Statistical Analysis

food samples were packed and shipped at ambient temperature. Upon receipt, samples were held by the collaborating laboratory at room temperature (24 ± 2°C). In addition to each of the test portions and the total plate count replicate, collaborators also received a test portion for each matrix labeled as “temperature control.” Participants were instructed to record the temperature of this portion upon receipt of the shipment, document results on the Sample Receipt Confirmation form provided, and fax to the Study Director. For both matrices, several shipments were delayed from getting to the testing facilities on time due to inclement weather. Upon receiving their packages on either Saturday or Monday, the testing laboratories were instructed to document the temperature of the samples and to continue testing. No laboratories were solely excluded from testing due to the delay in package receipt. Collaborators followed the appropriate preparation and analysis protocol according to the method specified for each matrix. For both matrices, each collaborator received 72 test portions of each food product (12 high, 12 low, and 12 controls for each method). For the analysis of the raw ground beef test portions by the 3M Petrifilm SALX System, a 25 g portion was enriched with 225 mL of prewarmed (41.5±1°C) 3M Salmonella Enrichment Base containing 3M Salmonella Enrichment Supplement (50 mg/L), homogenized for 2 min, and incubated for 18–24 h at 41.5±1°C. For the dry dog food test portions analyzed by the 3M Petrifilm SALX System, a 375 g portion was enriched with 3375 mL prewarmed (41.5±1°C) 3M Salmonella Enrichment Base containing 3M Salmonella Enrichment Supplement (50 mg/L), homogenized for 2 min, and incubated for 18–24 h at 41.5±1°C. Following enrichment of raw ground beef samples, the enrichment protocol for high microbial load foods was followed where a 0.1 mL aliquot of each test portion was transferred into 10.0 mL Rappaport-Vassiliadis R10 (R-V R10) broth and incubated for 8–24 h at 41.5±1°C. After incubation, a loopful of the secondary enrichment was streaked directly onto hydrated 3M Petrifilm SALX Plates and incubated for 24 ± 2 h at 41.5±1°C. For dry dog food samples, the enrichment protocol for low microbial load foods was followed where a loopful of the primary enrichment was streaked directly onto hydrated 3M Petrifilm SALX Plates and incubated for 24±2 h at 41.5±1°C. For both matrices, the 3M Petrifilm SALX Plates were examined for typical colonies (red to brown colony with a yellow zone or associated gas bubble, or both). Typical colonies were circled on the plate top film using a fine tip permanent black marker. The top of the 3M Petrifilm SALX Plate was lifted and a 3M Petrifilm SALX Confirmation Disk was placed onto the gel. The film was lowered and air bubbles were removed using a sweeping motion. The plates were incubated for 4–5 h at 41.5±1°C. After incubation, the circled colonies were observed for color change: red/brown to green blue, blue, dark blue, or black. Typical colonies were transferred to triple sugar iron/lysine iron agar (TSI/LIA) slants and confirmed following the standard reference methods. Additionally, for each matrix analyzed by the 3M Petrifilm SALX System, aliquots of the primary enrichment were transferred to the secondary enrichments and confirmed following procedures outlined in the MLG or BAM. Test Portion Analysis

AOAC Official Method 2014.01 Salmonella in Selected Foods 3M ™ Petrifilm ™ Salmonella Express System First Action 2014

[Applicable to detection of Salmonella spp. in raw ground beef (25 g), raw ground chicken (25 g), pasteurized liquid whole egg (100 g), raw ground pork (25 g), cooked chicken nuggets (325 g), frozen uncooked shrimp (25 g), fresh bunched spinach (25 g), dry dog food (375 g), and stainless steel. Not applicable to some lactose-positive Salmonella species.] See Tables 2014.01A and B for results of the interlaboratory study supporting acceptance of the method. See Appendix available on the J. AOAC Int . website for detailed tables of results of the collaborative study. Caution: Do not use the 3M Petrifilm SALX System method

in the diagnosis of conditions in humans or animals. To reduce the risks associated with exposure to chemicals and biohazards, perform pathogen testing in a properly equipped laboratory under the control of trained personnel. Always follow